2-hydroxy-2-carboxyethyl-n-(7- or 8-chloro-4-quinolinyl)anthranilate

ABSTRACT

2-Hydroxy-2-carboxyethyl-N-(7- or 8-chloro-4quinolinyl)anthranilate. The compounds are useful as analgesic agents.

United States Patent [1 1 Theriault et al. Oct. 9, 1973 Z-HYDROXY-Z-CARBOXYETHYL-N-(7- OR [58] Field of Search 260/287 R 8-CHLORO-4- QUINOLINYL)ANTHRANILATE [56] References Cited [75] Inventors: Robert John Theriault, Kenosha, UNITED STATES PATENTS v Wis.; Earl Elmer Fager, Lake Villa; 3,479,360 11/1969 Allais 260/287 R Norman Earl Wideburg, Waukegan, 3,679,687 7/1972 Wasley l 260/287 R both of lll. Primary Examiner-Donald G. Daus [73] Ass1 nee: Abbott Laboratories, North g Chicago, In. Attorney-Robert L. Nlblack [22] Filed: Apr. 13, 1972 57 ABSTRACT [2l] App]. No.: 246,091 2-Hydroxy-2-carboxyethyl-N-(7- or 8-chloro-4- quinolinyl)anthranilate. The compounds are useful as analgesic agents.

3 Claims, No Drawings 1 2-HYDROXY-Z-CARBOXYETHYL-Na(7-0R 8-CHLOR04 QUINOLINYUANTHRA'NIILATE I DETAILED DESCRIPTION OF THE INVENTION This invention relates to derivatives of 2,3-dihydroxypropyl-N-('7- or 8"-chloro-4'- quinolinyl)anthranilate and to processes for preparing and using the compounds.

The compounds of this invention, 2'-hydroxy-2'- carboxyethyl-N-(7-chloro-4-quinolinyl)anthranilate and 2-hydroXy-Z-carboXyethyI-N-( 8-chloro-4- quinolinyl)anthranilate are repersented by generalFormula I cocmcnon- A wherein the chloro is in the 7- or 8-position:

The compounds of this invention are mild, aspirinlike analgesic agents when administered orally towarm-blooded mammals in dosages of from 200-300 mg./kg. daily, preferably in divided doses, e.g., four to six times daily. The analgesic activity was initially established in the acetic acid writhing assay (Whittle, Brit. J. PharmcoL, 22: 246 [1964]).

Marasmius rotula SS-9. NRRL-3944 and cultures belonging to the Class: Shizomycetes I Order: Actinomycetales Family: Streptomycetaceae Genus: Streptomyces Species: Streptomyces sp. Act-9 NRRL3947 The compounds can be prepared by subjecting the appropriate 7- or 8-chloro compound ofFormulaII to microbial transformation according to the'following reaction sequence:

NRRL 3306 or NRRL 3944 or NRRL 32047 wherein the chloro is in the 7- or 8-position. The synthesis of the compounds of Formula II, 2,3- dihydroxypropyl-N-(7-chloro-4- quinolinyl)anthranilate and 2,3-dihydroxypropyl-N-(8- chloro-4-quinolinyl)anthranilate, are described'in Belgian Pat. Nos. 678,551, published Sept.' 28, I966 which Generally speaking, the compounds of this invention are prepared by inoculating a culture of one of the above-described microorganisms from 14 day old agar slant cultures into multiple flasks containing a suitable media. The inoculated flasks are incubated on ashaker for from 48-96 hours, at which time. 0.050.l percent of the powdered substrate, 2,3-dihydroxypropyl-N-('7- or 8-chloro-4-quinolinyl )anthranilate (50- 'mg-./ 1 00 ml. medium) is added to each flask. After 'addition'of the substrate, the flasks are returned to the shaker and harvested at the optimum times of 336-408 hours.

The harvested fermentation flasks are pooled and the beer is adjusted to pH 9.0 with NI-I OH. The fermentation'beer is then extracted twice with 2 volumes of the ethyl acetate and the extract is evaporated to dryness in vacuo. The ethyl acetate residue is then dissolved in a suitable solvent such as methanol or ethanol, and the resulting product obtained by either column or thinlayer chromatography (TLC).

The following fermentation media are used in the practice of this invention:

FERMENTATION MEDIA A Grams/liter Glucose monohydrate (added post sterlization) 50.0 Soy fluff flour 5.0 Yeast extract 5.0 KH PO 2.3 K HPO 0.84

The pH is adjusted to approximately 6.5 and deionized water is added to adjust the volume to 1.0 liter.

The pH is adjusted to 6.87.0 and deionized water is added to adjust the volume to 1.0 liter.

tion.

EXAMPLE 1 Preparation of 2-hydroxy,-2-carboxyethyl-N'-(7-chloro- 4-quinolinyl)anthranilate Marasmius rotula SS-9 NRRL-3944 was inoculated from 14 day old agar slant cultures into a series (approximately 200) of sterile cotton plugged 500 ml. Erlenmeyer flasks containing 100 ml. of medium A. The inoculated flasks were incubated on a Gump rotary shaker (250 rpms) at 28C. After 72 hours incubation, 0.05 percent of 2,3-dihydroxypropyl-N-(7-chloro-4- quinolinyl)anthranilate (50 mg./l00 ml. medium) was added in powdered form to each flask. The flasks were again incubated on the shaker and were sampled at various ages during the fermentation, as previously described, for TLC analysis-in order to determine the op- The following examples further illustrate this inventimal harvest age. All flasks were harvested at the peak yield of the major microbial conversion product. The contents (whole culture) of each flask were pooled and. filtered. The mycelia was then extracted with acetone: 0.01M NH OH (1:1). The mycelial aqueousacetone extract was adjusted to pH 7'.O with HCl, combined with the filtered beer and concentrated to'one-tenth quinolinyl)anthranilate or volume under vacuum. The concentrated, filtered beer and mycelial extract werethen-passed over a Darco' 6-60 carbonzcelite 545 111) column to absorb the desired product. The column was washed with water, chloroform and'finall'y 50 percent'aqueous acetone. The rnicrobialtransformation product was eluted. with the genus Streptomyces were inoculated into identical flasks containing 100 mleof medium E. Afterinoculation, the flasks were incubated at 28C. on a Gump rotary shaker (250 rpms) for 48 hours to 72hours. At

that time, growth was sufficient to permit the addition of 0.05 percent of 2,3-dihydroxypropyl-N-(7 chloro-4- 0.05 percent -of {2,3- dihydroxy-propyl-N-( 8.-chloro-4-quinolinyl =anthr-anilate ('50 mgJ IOO ml. medium) in powdered form. The flasks were again incubated on thefl shaker. and were sampled and extracted for thin-layer chroma- I tography analysis as. previously described.

0.1M NH OH:acetone (1:1 The eluate fractions were pooled and driedi'n vacuopand the. residue was ex tracte'd with ethanol: 0.25N Nl-LOH (10018). This extract was diluted with one volume of ethyl acetate and applied to a silica gel GF-254 (Merck-Darmstadt) 7 column slurry packed with. ethyl acetate:

ethanol:0.25N Nl-LOH (50:50:4 The column was developed with the same solvent system. Eluate fractions tomyces' species ACT-9 NRRL--'3947 each produce a ,mic'robial transformation "product from the 7+chl'oro substrate which ha-s the same TLC; R UV adsorption at '254 millimicronsandfluorescence at=336 millimi- 5 crons as does theproduct produced byMarasr'nius rotula SS-9 NRRL-*3944 described in Example 1. These three cultures transform the 8-chloro substrate:

' to yield products having TLC, R, and UV radiation containing the desired conversion product were combined and dried under vacuum. The residue was extracted with coldethanol, filtered and further purified by preparative thin-layer chromatography on a series of 20 X 20 cm glass plates coated with silica gel GF (Merck-Darmstadt) about 500 microns in thickness.

The plates were developed in a solve'nt. system consisting of CH Cl iCH oHzNl-hOl-l (75:25:11. The desired product waseluted with acidified 90 percent-aqueous acetone, and further purified by diffejre'ntial solventzex V chloro substrate indicating the same characteristics identicalwith the products from the 7- microbial t'ransfoiy mation of either substrate. I g I The compunds of this inventioncan be formulated into various pharmaceutical dosage forms such as tabl'ets,.capsules; pills,'-steril-e aqueous or non-aqueous so lut'ions'for'parenteral administration and the like,-for

immediate or sustained release, by combining one or traction of the dried residue, The product is repre ff sented by the formula This structure was confirmed by 100 MH nmr spectroscopy and by high resolution mass spectroscopy. Exact mass measurement: 386.0667 Calculated for C, H, N O Cl: 386.0669 The product does not have a' sharp melting point but softens beginning at 142C.

EXAMPLE 2 Preparation of 2-hydroxy-2-carboxyethyl-N-(7- and 8- chloro-4-quinolinyl)anthranilate Cultures belonging to the genera Coprinus and Mara Marasmius were inoculated from 14 day old agar slant cultures into sterile plugged 500 ml. Erlenmeyer flasks containing 100 ml. of medium A. Cultures belonging to more of the active compounds with .one orimorepharjm aceu'tica'lIy acceptable carriers or diluents. according to methods well known in the art. Such dosage forms may additionally.includeexcipients; binders, fillers,v fla 1 i, A 'voringfland sweetening agents and otehr therapeutically I ll COCHzCHOHCOH wherein the chloro atom is in the 7- or 8-position.

2. A compound of claim 1: 2-hydroxy-2-carboxyethyl-N-( 7-chloro-4-quinolinyl )anthranilate 3.'A compound of claim 1: 2-hydroxy-2-carboxyethyl- N-( 8-chloro-4-quinolinyl)anthranilate. 

2. A compound of claim 1: 2-hydroxy-2-carboxy-ethyl-N-(7-chloro-4-quinolinyl)anthranilate.
 3. A compound of claim 1: 2-hydroxy-2-carboxy-ethyl-N-(8-chloro-4-quinolinyl)anthranilate. 